The present invention relates to novel formulations comprising lipoic acid and the benzyl ester of hyaluronic acid with an esterification degree ranging between 75 and 100% for use in the repair of chronic ulcers, the regeneration of damaged skin, the prevention of skin aging and, when administered by the systemic route, to treat peripheral neuropathies and intoxications by chemical and/or biological agents.
Lipoic acid, also known as α-lipoic acid or thioctic acid, is a molecule physiologically produced by human body, in which it plays a role as co-factor in important enzymatic reactions in various metabolic processes, including Kreb's cycle. Due to its chemical structure, lipoic acid is prone to oxidation-reduction reactions, giving its reduced form of dihydrolipoic acid. It is also able to act as a carrier for electrons or acetyl groups or other acyls. Lipoic acid has some special features that make it not only extraordinarily effective as an antioxidant but also absolutely indispensable to human body in fighting the damage associated with free radicals formation. These features are:
Due to these properties (see also:
It has been demonstrated that lipoic acid administered by either the oral or parenteral route is effective in protecting the skin from inflammation (
The present invention relates to novel formulations of lipoic acid associated with a polysaccharide which remarkably improves its performance in terms of distribution homogeneity and, thanks to its peculiar features, acts synergically with lipoic acid, enhancing its antioxidant, anti-inflammatory, free-radical-scavenging and wound healing effects. Thus, the limitations of the current state of the art, where lipoic acid is simply vehicled in pharmaceutical/cosmetic carriers, are overcome. According to the invention, the polysaccharide associated with lipoic acid is hyaluronic acid (HA), known for its ability to act as a delivery system for the release of pharmacologically and/or biologically active substances to be administered by the topical route (
HA is a heteropolysaccharide composed of alternating residues of D-glucuronic acid and N-acetyl-D-glucosamine. It is a linear-chained polymer with a molecular weight that varies between 50,000 and 13 x 106 Da, according to the source from which it is obtained and the methods used to prepare it. HA is ubiquitous in nature (for example in the pericellular gels, the fundamental substance of connective tissue in vertebrate organisms, in the synovial fluid of joints, the vitreous humor, the umbilical cord) and exercises numerous important functions in the biological organism. It acts as a mechanical support for the cells of many tissues, such as the skin, the tendons, the muscles and the cartilage, and by means of its membrane receptor CD44, it modulates many processes involved in cell physiology and biology, such as cell proliferation, migration and differentiation, and angiogenesis. It has also been demonstrated that HA plays a fundamental role in the tissue repair process, both from a structural point of view (in organising the extracellular matrix and regulating its hydration) and as stimulator of a vast series of processes in which it intervenes either directly or indirectly (clot formation, phagocyte activity, fibroblast proliferation, neovascularisation, re-epithelialisation, etc.) (
Also known is the use of HA derivatives in the formation of fibres (
The hyaluronic acid used in the present invention may be obtained from any source, for example by extraction from rooster combs (
The present invention relates to formulations of lipoic acid associated with the benzyl ester of hyaluronic acid with an esterification degree ranging between 75 and 100% to be used substantially for the topical treatment of all skin conditions characterised by oxidative stress and the massive presence of free radicals, such as autoimmune diseases, or irritative, atopic, contact or allergic dermatitis, inflammatory states, wounds, or ulcers. The formulations of the present invention are also suitable for the dermocosmetic treatment of the signs of aging and photoaging of the skin. In the present invention, lipoic acid used at concentrations ranging between 0.5 and 15%, and preferably between 1 and 10%, has been successfully associated with derivatives of hyaluronic acid obtained by chemical modification by esterification with alcohols of the aliphatic, araliphatic, cycloaliphatic, aromatic, cyclic and heterocyclic series (HYAFF®).
The derivatives obtained as described herein maintain the biological properties of biocompatibility and biodegradability of the starting molecule, but they give a better mechanical performance, with better bioadhesiveness, a controlled release of the lipoic acid and a residence time of the final association of HA/lipoic acid that significantly enhances all the chemical-physical, biological and pharmacological properties of both hyaluronic acid and lipoic acid.
Thanks to their special features, the formulations described herein can also be used for the systemic and/or locoregional administration, for example in treatment of intoxication by chemical and/or biological agents and in peripheral neuropathies. Of the various conditions that can benefit from topical administration of the formulations described herein, particularly important are venous ulcers. Venous ulcers of the lower limbs have considerable social impact because of their incidence in the population (about 1%) and slow rate of healing. Generally, repair of chronic ulcers is seriously hampered by the disregulating effect of chronic inflammation on both the cytokine cascade and the physiological repair processes. In particular, venous ulcers are characterised by overexpression of metalloprotease (MMP) which, together with the disappearance of their physiological tissue inhibitors, determine excessive proteolysis of the extracellular matrix (ECM). Such pathological proteolysis is determined by the presence, in the tissues surrounding the ulcer, of a considerable quantity of iron which induces the formation of free radicals and the recruitment of macrophages which undergo oxidative stress as a result of excessive phagocytosis of the metal. The abundance of iron activates MMP expression as described above, and MMP activity is further amplified by feedback established by the ECM degradation products themselves. The problem of healing venous ulcers is therefore solved by the formulations that are the subject of the present invention, constituted by lipoic acid and hyaluronic acid and/or the derivatives thereof. Lipoic acid, being an antioxidant and iron chelator, takes the free radicals, especially iron, from the tissues surrounding the ulcer, hindering the action of MMP and thus favouring the formation of ECM. Lipoic acid is aided in this by the presence of hyaluronic acid and/or the derivatives thereof. Of the numerous derivatives, particularly useful for this type of application is the benzyl ester of hyaluronic acid (HYAFF®-11) in which the percentage of esterification ranges from 0.1 to 100%, preferably between 50 and 100% and more preferably still between 75 and 100%. Worked in the form of a non-woven fabric, it forms a soft pad that can be adapted to fit the shape and size of the ulcer, and is also bioadhesive. Its hyaluronic acid content is active at various levels:
The synergic action of the formulations of the present invention causes faster, more effective healing of the ulcer than can be achieved with the dressings available to date. New therapeutic possibilities are therefore opened up that go beyond the current state of the art.
Similar results are achieved by using, according to the type of lesion to be treated, hyaluronic acid and/or the derivatives thereof in the form of a gel, cream, ointment, lotion, suspension for intradermal/subcutaneous use (including fillers) for the treatment of skin disorders such as dermatitis of various kinds and autoimmune diseases, for the prevention and dermocosmetic cure of the signs of aging and photoaging and for all skin manifestations characterised by a non-physiological presence of free radicals and alterations in the oxidation-reduction balance. In such situations the antioxidant effect and free-radical-scavenger effect of lipoic acid are enhanced and amplified by the presence of HA and/or the derivatives thereof. Moreover, hyaluronic acid ensures an even distribution of lipoic acid within the formulation thus guaranteeing its constant and continuous release once administered.
Therefore, the invention also relates to to formulations of lipoic acid associated with the benzyl ester of hyaluronic acid with an esterification degree ranging between 75 and 100% for the systemic administration such as oral, rectal, locoregional, injective (intradermal, subcutaneous, intramuscular, intravenous, intraperitoneal) routes,for use in the treatment of other disorders that benefit from free radical scavenging, such asintoxication by chemical and/or biological agents: besides the effects described previously, HA, rapidly captured by the liver cells, enables the lipoic acid to act specifically on the liver cells by interacting with its membrane receptor CD44, thus stimulating their regeneration;
peripheral neuropathies: HA contributes significantly to a profile of smooth, constant and continuous release.
For purely descriptive purposes, and without being limited by the same, The present invention is disclosed in more detail in the following examples.
A non-woven fabric of HYAFF® 11-p75 measuring $10X10 cm is soaked with a 5% solution of lipoic acid in absolute ethanol. The treatment involves leaving the non-woven fabric in the ethanol solution of lipoic acid for 3 minutes. At the end of this time, the excess liquid is discarded and the non-woven fabric is dried by eliminating the residue solvent by vacuum extraction on a hotplate heated to 30°C. The product is then analysed to determine the quantity of lipoic acid that has been absorbed by the non-woven fabric, which should be between 100 and 200 mg for each piece measuring 10 X 10 cm.
Examples 2-8 are not part of the invention.
The percarboxylated derivative is solubilised in DMF (dimethylformamide) at a concentration of 80-120 mg/ml, and coagulated in a solution constituted by 2% lipoic acid in absolute ethanol. This treatment enables the lipoic acid to be absorbed by the percarboxylated derivative. The product, in the form of a film, is dried in a vacuum on a hotplate heated to 30°C, and analysed to determine the quantity of lipoic acid it contains, which should be between 100 and 200 mg for each piece measuring 10 X 10 cm.
5 grams of sulphated hyaluronic acid with a degree of sulphation of 3 (7.05 millimole) and a mean molecular weight of 200 Kda is solubilised in 50 ml of water. The solution is vigorously stirred with a magnetic stirrer while slowly adding a solution of 1% lipoic acid in ethanol, prepared by dissolving 500 mg of lipoic acid in 50 ml of 96% ethanol.
The resulting emulsion is freeze-dried.
The product thus obtained is analysed to determine the quantity of lipoic acid contained in the freeze-dried product, which should be between 80 and 100 mg of lipoic acid per gram of sulphated derivative.
2 grams of dodecyl amide of hyaluronic acid (HYADD™ -3) at 5% (4.88 millimole) with a mean molecular weight of 200 Kda is solubilised in 100 ml of water. The solution is vigorously stirred with a magnetic stirrer while slowly adding a solution of 20% lipoic acid in 96% ethanol, prepared by dissolving 200 mg of lipoic acid in 1 ml of 96% ethanol. The resulting emulsion is transferred in a 2-ml glass syringe and steam-sterilised. A topical dressing in the form of a sterile gel comprising an amide derivative containing 10% lipoic acid is thus obtained. The product is analysed to determine the quantity of lipoic acid contained in the gel, which should be between 80 and 100 mg of lipoic acid per gram of amide derivative of hyaluronic acid.
2 grams of hexadecyl amide of hyaluronic acid (HYADD™-4) at 3% (4.90 millimole) with a mean molecular weight of 500 Kda is solubilised in 100 ml of water. The solution is vigorously stirred with a magnetic stirrer while slowly adding a solution of 10% lipoic acid in 96% ethanol, prepared by dissolving 100 mg of lipoic acid in 1 ml of 96% ethanol. The resulting emulsion is transferred into a 2-ml glass syringe and steam-sterilised. A topical dressing in the form of a sterile gel comprising an amide derivative of hyaluronic acid containing 5% lipoic acid is thus obtained. The product is analysed to determine the quantity of lipoic acid container in the gel, which should be between 40 and 50 mg of lipoic acid per gram of amide derivative of hyaluronic acid.
1 gram of hyaluronic acid cross-linked to a degree of 10% (2.52 millimole) with a mean molecular weight of 200 Kda is solubilised in 100 ml of water. The solution is vigorously stirred with a magnetic stirrer while slowly adding a solution of 10% lipoic acid in 96% ethanol, prepared by dissolving 100 milligrams of lipoic acid in 1 ml of 96% ethanol. The resulting emulsion is transferred to a 2-ml glass syringe and steam-sterilised. A topical dressing in the form of a sterile gel comprising an auto-cross-linked derivative of hyaluronic acid containing 10% lipoic acid is thus obtained.
The product is analysed to determine the quantity of lipoic acid contained in the gel, which should be between 80 and 100 mg of lipoic acid per gram of cross-linked derivative of hyaluronic acid.
A prefilled syringe of 2 ml contains:
The resulting suspension is suitable for injection and packed in prefilled syringes and steam-sterilised.
A prefilled syringe of 2 ml contains:
The resulting suspension is suitable for injection and packed in prefilled syringes and steam-sterilised.
This list of references cited by the applicant is for the reader's convenience only. It does not form part of the European patent document. Even though great care has been taken in compiling the references, errors or omissions cannot be excluded and the EPO disclaims all liability in this regard.